Gene expression analysis of diet induced obesity model zebrafish
ABSTRACT: As microarray based gene expression profiling is well suited to study the complex diseases such as obesity, we revealed gene expression changes of fat tissues on obesity model zebrafish to elcidate the pathophysiological function of each fat tissue in metabolic syndrome. Zebrafish in over-feeding group were fed three times per day with Artemia (60 mg cysts/fish/day) through 8weeks. 1week over-feeding group were fed three times per day with Artemia (60 mg cysts/fish/day) through 1week. For caloric restriction, zebrafish were fed with Artemia (2.5 mg cysts/fish/day) for 2 weeks after over-fed with Artemia for 8 weeks.
Project description:As microarray based gene expression profiling is well suited to study the complex diseases such as obesity, we revealed gene expression changes of fat tissues on obesity model zebrafish to elcidate the pathophysiological function of each fat tissue in metabolic syndrome. Overall design: Zebrafish in over-feeding group were fed three times per day with Artemia (60 mg cysts/fish/day) through 8weeks. 1week over-feeding group were fed three times per day with Artemia (60 mg cysts/fish/day) through 1week. For caloric restriction, zebrafish were fed with Artemia (2.5 mg cysts/fish/day) for 2 weeks after over-fed with Artemia for 8 weeks.
Project description:We used a Diet-Induced-Obesity approach using the zebrafish (Danio rerio) based on overfeeding to analyze the liver transcriptomic modulation in the disease and to determine how the obesity affects the immune response against an acute inflammatory stimulus as the lipopolysacharide (LPS). Overfed zebrafish were obese and showed signs of steatosis in their liver. Furthermore, the gene modulation profile resembled to that observed in humans. After the inflammatory stimulus, there was a clear differential immune response in normal and overfed fish. In normal fish, the response to LPS reported a typical host defensive reaction comparable to the one occurring in stimulated mammals whereas there was not any significantly modulated gene when comparing the expression of liver from LPS-stimulated with non-stimulated obese zebrafish. Overall design: Healthy fish were fed two times a day, one with Artemia and the other with standard fish chow while obese fish were fed fish chow two more times a day during 8 months. 10 μg of pure LPS (Sigma) or PBS was intraperitoneally injected and livers were sampled and pooled at 3 hours post injection.
Project description:In this study, microarrays were used to investigate the larval cod transcriptome response to zooplankton supplementation in the diet. Overall design: The experiment consisted of three different feeding regimes/diets as follows: Treatment 1 (6 replicate tanks): rotifers/Artemia with Ori-Green (Skretting, USA) enrichment (RA) 3 feedings per day (9 am, 3 pm and 9 pm), with rotifers fed until the fish reached ~ 9 mm [~ 30 days post-hatch (dph)]. Treatment 2 (4 replicate tanks): rotifers with Ori-Green enrichment, supplemented with 5-10% wild caught zooplankton (RA-Zoo: zooplankton were > 90% copepods) from 2 dph until 30 dph. Treatment 3 (6 replicate tanks): Rotifers/Artemia with Ori-Green enrichment 4 days per week and Rotifers/Artemia with protein hydrolysate (RA-PH) enrichment 3 days per week (Monday, Wednesday and Friday),with rotifers fed until the fish reached ~ 9 mm. Whole larvae from each feeding regimes/diets were sampled when fish in particular treatment reached an average length of ~ 9 mm. Individual DNAse-treated, column-purified total RNA samples from 8 fish per group (9 mm RA, 9 mm RA-PH and 9 mm RA-Zoo larvae) were used for microarray analysis using the Atlantic cod 20K oligonucleotide microarray platform (Booman et al. 2011) and a common reference experimental design. On each array, RNA from an individual fish was hybridized together with a common reference RNA. All test samples were labeled with Cy5. The common reference was labeled with Cy3.
Project description:Silver nanoparticles (NPs) are extensively used due to their antimicrobial activity and, therefore, their input into the ecosystem will increase. Silver can be bioaccumulated by low trophic level organisms and, then, incorporated into the food chain, reaching high level predators. The objectives of this study were to test the acute toxicity of N-vynil-2-pirrolidone/polyethylenimine (PVP-PEI) coated Ag NPs of 5 nm to brine shrimp (Artemia sp) larvae and to assess bioaccumulation and effects of silver transferred by the diet. For the later, brine shrimps were exposed to two different concentrations of Ag NPs, 100 ng/L as an environmentally relevant concentration and 100 µg/L as a likely effective concentration, in parallel with an unexposed control group and, then, used to feed zebrafish during 21 days in order to simulate two trophic levels of a simplified food web. For brine shrimp larvae, EC50 values ranged from 7.39 mg Ag/L (48 h post hatch larvae (hph) exposed for 48 h) to 19.63 mg Ag/L (24 hph larvae exposed for 24 h. Silver accumulation was measured in brine shrimps exposed to 0.1 and 1 mg/L of Ag NPs for 24 h. In zebrafish fed with brine shrimps exposed to Ag NPs, intestine showed higher metal accumulation than liver, although both organs presented the same pattern of dose and time-dependent metal accumulation as revealed by autometallography. Feeding of zebrafish for 3 days with brine shrimps exposed to 100 ng/L of Ag NPs was enough to impair fish health as reflected by the significant reduction of the lysosomal membrane stability and the presence of several histopathological conditions in the liver. Overall, results showed that Ag NPs were able to exert toxic effects on zebrafish through dietary exposure, even at an environmentally relevant concentration, which should act as concern of the need of studies in further detail about real impact of nanomaterials in the environment. Overall design: 20 samples per group, 5 replicates (4 liver or 4 intestine per replicate), 2 groups x 2 times, control and C1 (0.1 mg/L) of Ag NPs, 3 and 21 days of exposure
Project description:Dieldrin is a legacy pesticide that has multiple modes of action (MOA) that include being an estrogen receptor agonist, GABA receptor antagonist, and a chemical that disrupts mitochondrial function. There is also evidence that dieldrin exposure is significantly associated with an increased risk for neurodegeneration in humans. The objective of this thesis was to clarify the effects of dieldrin in the hypothalamus, the major neuroendocrine region of the brain, in the zebrafish (Danio rerio). Zebrafish were fed pellets containing 0.03, 0.15, or 1.8 µg/g dieldrin for 21 days and a global gene expression analysis was performed to characterize cellular processes and pathways affected by dieldrin. Overall design: Reproductive adult zebrafish (4-6 months of age) were obtained from Mirdo Inc. (Montréal, QC, Canada). Zebrafish were housed at the Canadian Rivers Institute (Saint John, NB, Canada) in 20 L tanks. After an acclimation period of 3 weeks, zebrafish were divided into 9 tanks per experimental group giving a total of 36 tanks, each with active carbon within the tank as an extra precaution to minimize any exposure to water-borne dieldrin. Five fish were added to each tank, with the goal of maintaining a 3 to 2 female to male sex ratio. Zebrafish in designated tanks were fed 0.065 g of the control or one of the three contaminated feeds twice a day representing a daily intake of 0.13 g (approximately 4% of fish body weight per tank). Feeding took place at 10 am and 4 pm each day.
Project description:The effect of a year-long 10 reduction in water temperature on global gene expression in tail skeletal muscle from adult, male zebrafish was determined using a long oligonucleotide probe set (16,399 65mers from Compugen) spotted onto glass slides. Outbred male zebrafish were obtained from a commercial supplier (Liles Tropical Fish, FL) at 6 months of age. Fish were maintained at 28 until 10 months of age. (see Gerhard et al., Exp Gerontol 37,1055-68, 2002) For temperature reduction, water temperature was decreased by 2.5 per week for 4 weeks. Fish were maintained until 22 months of age. Fish were fed fish flakes (Wardely Corp. Secaucus, NJ) twice per day. Each feeding was a discrete event in which a small measured amount of granular food was sprinkled on the water surface. A second small aliquot was offered if the first has been eaten within a few minutes. The feeding stopped when food from the previous aliquot has not been eaten and feeding behavior has ceased. By providing a consecutive series of small aliquots until food is no longer accepted, every member of the tank has an opportunity to eat until satiety, yet the amount of uneaten food is minimized. At 22 months of age, fish were euthanized by decapitation. Total RNA was harvested from a pool of tail muscle samples from 5 fish. Two independent pools of 5 fish per pool were collected from fish maintained at 18 and 28. Flip-dye hybridizations were performed on each pool for a total of 4 hybridizations in this study. Experiments DAR011d0001 and DAR011d0002 are flip-dye hybridizations from Pool 1. Experiments DAR011d0003 and DAR011d004 are flip-dye hybridizations from Pool 2.
Project description:We identified that the adiponectin gene expression in rainbow trout muscle decreased by restrected feeding. In order to identify the genes differently expressed by the same treatment, micrarray analysis was conducted Fish were fed ad libitum once a week (RF, restricted feed group) or fed ad libitum twice per day (control). After 1 month, the muscle was desected from 4 individuals from each group.
Project description:Recent evidence has suggested that fluoxetine, a serotonin-reuptake inhibitor and emerging environmental contaminant, can have non-targeted effects on metabolism in fish exposed to this waterborne pollutant. Using the highest, environmentally relevant, detectable level of fluoxetine (540 ng/L) we examined the impact of fluoxetine on the miRNA profile in the liver of zebrafish that were both fed and fasted for a period of 7 days. These results were further compared to the miRNA profile of zebrafish fasted and fed for 7 days, which were not exposed to fluoxetine. Results indicated that several miRNA that were involved with downregulating genes/pathways in response to fasting were also upregulated in fish exposed to fluoxetine, irrespective to fasting or feeding. These results suggest fluoxetine can have non-targeted effects on metabolic pathways mediated through miRNA expression. Furthermore, specific miRNA (dre-let-7d & dre-miR-140-5p) were found to target the catalytic subunit (AMPKa1 & AMPKa2, respectively) of AMP-Kinase, a master regulator of metabolism. Using predictive software and qPCR validation, combined with the expression profile of these two miRNA, we were able to establish a significant relationship between the expression of these specific miRNA to the downregulation of AMPKa subunit under the influence of 540 ng/L fluoxetine. Adult, female zebrafish were either fed or fasted for 7 days with and without the presense of 540 ng/L fluoxetine, and livers extracted and miRNA purified for miRNA microaary experiment.
Project description:The aim of this experiment was to explore transcriptomic changes in the distal gut of Altantic salmon fed five experimental and two control diets. The experimental diets were isonitrogenous, isolipidic and isocaloric. They had similar content of fish meal (~22%) and similar total plant protein content (~45%), but differed in the contents of soya protein concentrate (SPC) and bean protein concentrate (BPC). These two ingredients were used to replace each other (either partially or fully) at the levels of incorporation ranging from 0 to 45%, with ~11% increments. As a result, the experimental diets contained either 45% SPC and 0% BPC (S45), 34% SPC and 11% BPC (S34B11), 22% SPC and 22% BPC (S22B22), 11% SPC and 34% BPC (S11B34) or 0% SPC and 45% BPC (B45). The S45 and B45 diets were single plant protein diets, while the others (S34B11, S22B22 and S11B34) were mixed plant protein diets. The control diets were enriched with fish meal (FM diet) or soybean meal (SBM diet) to provide negative and positive controls for gut inflammation (enteritis), respectively. The study was conducted at EWOS Innovation Research Facility in Dirdal (Norway) Atlantic salmon (Salmo salar L.) of the SalmoBreed strain were supplied as fertilized eggs and hatched on site. Mixed-sex juvenile salmon in groups of ~150 fish were transferred to 26 indoor tanks (0.6 x 0.6 x 0.6 m), with ~60 L of freshwater flowing at a rate of 3.9 L/min and continuous aeration. The temperature of water was regulated at 13°C and all animals were exposed to a constant light regime (24 h light/day). Fish were fed a commercial EWOS Start 1 diet and acclimated to the experimental conditions for 2 weeks, after which (at body mass ~1.5 g) they were subjected to a 56-day feeding trial. Tanks were randomly assigned to the dietary treatments, with 4 replicate tanks per each experimental diet (S45, S34B11, S22B22, S11B34 and B45) and 3 replicate tanks per each control diet (FM and SBM diets). Fish were fed to satiation prior to the feeding trial and during the dietary manipulation using automatic band feeders (Holland Technology, Norway). During the feeding trial, all fish in each tank had their biomass recorded on days 0 (start of experiment), 14, 28, 42 and 56 (end of experiment) for evaluation of growth performance.
Project description:Gaining understanding of common complex diseases and their treatments are the main drivers for life sciences. As we show here, comprehensive protein set analyses offer new opportunities to decipher functional molecular networks of diseases and assess the efficacy and side-effects of treatments in vivo. Using mass spectrometry, we quantitatively detected several thousands of proteins and observed significant changes in protein pathways that were (dys-) regulated in diet-induced obesity mice. Analysis of the expression and post-translational modifications of proteins in various peripheral metabolic target tissues including adipose, heart, and liver tissue generated functional insights in the regulation of cell and tissue homeostasis during high-fat diet feeding and medication with two antidiabetic compounds. Protein set analyses singled out pathways for functional characterization, and indicated, for example, early-on potential cardiovascular complication of the diabetes drug rosiglitazone. In vivo protein set detection can provide new avenues for monitoring complex disease processes, and for evaluating preclinical drug candidates. Male C57BL/6 mice (age 6 wks) were fed for 12 weeks with high-fat diet (HFD) and than distributed into 3 groups. Mice were than fed over 3 weeks with HFD without compound ('HFD'), HFD with 4 mg/kg/d rosiglitazone ('HFD_RSG') or with 100 mg/kg/d amorfrutin 1 ('HFD_A1'). In parallel, mice were fed for 15 weeks with low-fat diet (LFD) as healthy control subjects. In addition, a group of mice was treated with 37 mg/kg/d amorfrutin 1 during the whole 15 weeks of HFD feeding ('HFD+A1prev'). Finally, visceral white adipose tissue (WAT) was harvested and RNA extracted. --> 2-4 biological replicates (2 mice per each replicate, RNA of 2 mice always pooled).